Unraveling and resolving inefficient glucolipid biosurfactants production through quantitative multiomics analyses of Starmerella bombicola strains

Abstract

Glucolipids (GLs) are glycolipid biosurfactants with promising properties. These GLs are composed of glucose attached to a hydroxy fatty acid through a ω and/or ω‐1 glycosidic linkage. Up until today these interesting molecules could only be produced using an engineered Starmerella bombicola strain (∆ugtB1::URA3 G9) producing GLs instead of sophorolipids, albeit with a very low average productivity (0.01 g·L−1·h−1). In this study, we investigated the reason(s) for this via reverse‐transcription quantitative polymerase chain reaction and Liquid chromatography‐multireaction monitoring‐mass spectrometry. We found that all glycolipid biosynthetic genes and enzymes were downregulated in the ∆ugtB1 G9 strain in comparison to the wild type. The underlying reason for this downregulation was further investigated by performing quantitative metabolome comparison of the ∆ugtB1 G9 strain with the wild type and two other engineered strains also tinkered in their glycolipid biosynthetic gene cluster. This analysis revealed a clear distortion of the entire metabolism of the ∆ugtB1 G9 strain compared to all the other strains. Because the parental strain of the former was a spontaneous ∆ura3 mutant potentially containing other “hidden” mutations, a new GL production strain was generated based on a rationally engineered ∆ura3 mutant (PT36). Indeed, a 50‐fold GL productivity increase (0.51 g·L−1·h−1) was obtained with the new ∆ugtB1::URA3 PT36 strain compared with the G9‐based strain (0.01 g·L−1·h−1) in a 10 L bioreactor experiment, yielding 118 g/L GLs instead of 8.39 g/L. Purification was investigated and basic properties of the purified GLs were determined. This study forms the base for further development and optimization of S. bombicola as a production platform strain for (new) biochemicals

Ultrasound assisted in-situ separation of sophorolipids in multi-phase fermentation system to achieve efficient production of Candida bombicola

“…SLs has been regarded as a green and sustainable biosurfactants, but their high production costs limit largescale application and development. [21][22][23] Coupling fermentation on the basis of in-situ product separation is one of the most effective ways to reduce production costs and improve productivity (Table 2). Wang designed a simple in-situ separation device coupled with the bioreactor and adopted high cell density fermentation to achieve high-efficient production of SLs.…”

Microbial biosurfactant research: time to improve the rigour in the reporting of synthesis, functional characterization and process development

“…During process development, we recommend to generate highly pure standards of the biosurfactant mixture and then use these ‘standards’ (consisting of a mixture of congeners) to analyse broth samples containing the same mixture of biosurfactant congeners in the same ratio’s (Roelants et al ., 2016; Lodens et al ., 2020). Important to note is that one should carefully consider the ratio’s of the compounds/peaks in the samples versus the standards.…”

“…If these ratio’s remain the same this method is valid. Because the latter is not always the case, it is also recommended – if possible – to generate highly pure standards of the most abundant congeners available in the mixture, so these can be quantified (absolutely) separately, for example non‐acetylated C18:1 acidic sophorolipids, di‐acetylated C18:1 lactonic sophorolipids, non‐acetylated C18:1 acidic glucolipids (Roelants et al ., 2016; Lodens et al ., 2020). This will thus be a case to case issue, but the authors of this work active on these aspects, typically make sure both options are at hand.…”

Biosurfactants: Production and application prospects in the food industry

“…104,105,106 S. bombicola has a non-pathogenic nature and the capacity to produce large quantities of biosurfactant with a sophorolipid structure (around 177 g/L) from agricultural waste products and under conditions of a high concentration of hydrophobic compounds, making it promising for industrial applications as an emulsifying agent and stabilizer, especially in the food industry. 107,108,109 Yeasts of the genus Saccharomyces are also widely explored in the food area. Studies conducted as far back as the 1980s reported the use of intracellular compounds with emulsifying activity derived from yeasts of the genera Saccharomyces, such as S. cerevisiae, demonstrating the possibility of extracting mannoproteins in an easy manner from the cell wall that exhibit stability at different temperatures.…”

“…This yeast is considered to be a teleomorph of C. bombicola due to the high identity of the genetic material between the two (more than 98%) and its sexual reproduction stage 104,105,106 . S. bombicola has a non‐pathogenic nature and the capacity to produce large quantities of biosurfactant with a sophorolipid structure (around 177 g/L) from agricultural waste products and under conditions of a high concentration of hydrophobic compounds, making it promising for industrial applications as an emulsifying agent and stabilizer, especially in the food industry 107,108,109 …”